Response of Saccharomyces cerevisiae strains to antineoplastic agents

The effect of several antineoplastic agents on Saccharomyces cerevisiae strains has been investigated. Minimum inhibitory concentration (MIC), minimum cytotoxic concentration (MCC) and median effective concentration (EC₅₀) were determined to identify strains with inherent sensitivity to the agents tested. Several strains proved to be sensitive to the antimetabolites 5-fluorouracil and methotrexate as well as to doxorubicin and cis-platine. On the contrary m -amsacrine, procarbazine, vinca alcaloids, melphalan and hydroxyurea were inactive at concentrations up to 400 μg ml ⁻¹. The strain ATCC 2366, the most relatively sensitive to the agents tested, was used for studying the effect of treatment duration and of drug concentration on cell survival. Methotrexate and cis-platine, which according to MIC and MCC tests seemed ineffective for this strain, reduced survival significantly after 6 h of treatment. A correlation of the shape of the survival curves with MIC and MCC values was attempted.     

Effect of calcium channel blockers on the action of various antitumour agents in the yeast Saccharomyces cerevisiae

The cytotoxic effects of a 22 h treatment with four antineoplastic agents in the yeast Saccharomyces cerevisiae ATCC 2366 were investigated. Two agents, doxorubicin and 5-fluorouracil (5-FU), were effective in decreasing the colony-forming ability of yeast cells. Following examination under the light microscope, the effect of doxorubicin appeared to be, at least partially, due to killing of yeast cells whereas the effect of 5-FU was rather due to changes in cell structure leading to abnormal bud formation. For amsacrine (AMSA) and melphalan, cytotoxicity was totally absent. In the presence of diltiazem the above described effects were not significantly changed. When verapamil was added in the culture medium the cytotoxic activity of doxorubicin and 5-FU did not change. However, following treatment with AMSA in combination with verapamil, cell survival was significantly decreased whereas the presence of verapamil increased the yeast survival which was observed after melphalan treatment.     

A potassium transport mutant of Saccharomyces cerevisiae

A mutant in Saccharomyces cerevisiae required one hundred times more K⁺ than wild type for the same half maximal growth rate. Mutant cells and wild type cells grown at millimolar K⁺ did not show significant differences in Rb⁺ transport. In the mutant, a rapid K⁺ loss induced by azide or incubation (4 h) in K⁺-free medium decreased the Rb⁺ transport K _m by one half; in the wild type, those treatments decreased the Rb⁺ K _m twenty and one hundred times, respectively. Mutant and wild type did not show significant differences in Na⁺ transport and in the Na⁺ inhibition of Rb⁺ transport, either in normal-K⁺ cells or in K⁺-starved cells. The results suggest that either two systems or one system with two interacting sites mediate K⁺ transport in S. cerevisiae.Abbreviations YPD yeast-peptone-dextrose medium     

Cloning of the gene of Saccharomyces cerevisiae yeasts that determines resistance to the toxic action of cadmium ions

A gene conferring resistance to cadmium in Saccharomyces cerevisiae was isolated from a yeast gene library created on the basis of the pL3 vector. The phenotype of resistance is only expressed in the yeast cells with cloned DNA inserted into a multicopy plasmid. Integration of the plasmid into chromosome or introduction of the centromeric region into the plasmid decreases the level of cadmium resistance. The cloned Sau3A I fragment of the yeast chromosome is 3.5 kbp in size. Restriction analysis and subcloning experiments showed the gene to be located within 1.6 kbp of the XhoI-Sau3A I fragment of DNA. Instability was observed in the vicinity of the XhoI-Sau3A I fragment of the yeast DNA in Escherichia coli.     

Effects of tellurite on growth of Saccharomyces cerevisiae

The effects of potassium tellurite on growth and survival of rho⁺ and rho⁰ Saccharomyces cerevisiae strains were investigated. Both rho⁺ and rho⁰ strains grew on a fermentable carbon source with up to 1.2 mM K₂TeO₃, while rho⁺ yeast cells grown on a non-fermentable carbon source were inhibited at tellurite levels as low as 50 μM suggesting that this metalloid specifically inhibited mitochondrial functions. Growth of rho⁺ yeast cells in the presence of increasing amount of tellurite resulted in dose-dependent blackening of the culture, a phenomenon not observed with rho⁰ cultures. Transmission electron microscopy of S. cerevisiae rho⁺ cells grown in the presence of tellurite showed that blackening was likely due to elemental tellurium (Te⁰) that formed large deposits along the cell wall and small precipitates in both the cytoplasm and mitochondria.     

Effect of iron ions on the antioxidant enzyme activities in yeast Saccharomyces cerevisiae

A complex of physiological and biochemical indices has been compared in wild and isogenic catalase-deficient strains of Saccharomyces cerevisiae grown on the media with different iron ion concentrations is 2 times higher in cytosolic catalase deficient yeast. Superoxide dismutase activity grown in the medium with 500 microM of ferrous sulphate. Under such conditions, peroxisomal catalase deficient yeast had a 2-fold decreased activity of superoxide dismutase. There is a significant difference between TBA-reactive substances content of the wild and cytosolic catalase deficient strain. It has been suggested that the repletion of iron ions in the growth medium leads to the formation of lipid oxidation products. Catalase prevents TBA-reactive substances formation in the given conditions and plays a protective role.     

Involvement of an Actomyosin Contractile Ring in Saccharomyces cerevisiae Cytokinesis

In Saccharomyces cerevisiae, the mother cell and bud are connected by a narrow neck. The mechanism by which this neck is closed during cytokinesis has been unclear. Here we report on the role of a contractile actomyosin ring in this process. Myo1p (the only type II myosin in S. cerevisiae) forms a ring at the presumptive bud site shortly before bud emergence. Myo1p ring formation depends on the septins but not on F-actin, and preexisting Myo1p rings are stable when F-actin is depolymerized. The Myo1p ring remains in the mother–bud neck until the end of anaphase, when a ring of F-actin forms in association with it. The actomyosin ring then contracts to a point and disappears. In the absence of F-actin, the Myo1p ring does not contract. After ring contraction, cortical actin patches congregate at the mother–bud neck, and septum formation and cell separation rapidly ensue. Strains deleted for MYO1 are viable; they fail to form the actin ring but show apparently normal congregation of actin patches at the neck. Some myo1Δ strains divide nearly as efficiently as wild type; other myo1Δ strains divide less efficiently, but it is unclear whether the primary defect is in cytokinesis, septum formation, or cell separation. Even cells lacking F-actin can divide, although in this case division is considerably delayed. Thus, the contractile actomyosin ring is not essential for cytokinesis in S. cerevisiae. In its absence, cytokinesis can still be completed by a process (possibly localized cell–wall synthesis leading to septum formation) that appears to require septin function and to be facilitated by F-actin.     

Ketoconazole and miconazole alter potassium homeostasis in Saccharomyces cerevisiae

The effects of ketoconazole and miconazole uptake on K(+) transport and the internal pH of Saccharomyces cerevisiae were studied. The uptake of both drugs was very fast, linear with concentration and not dependent on glucose, indicating entrance by diffusion and concentrating inside. Low (5.0μM) to intermediate concentrations (40μM) of both drugs produced a glucose-dependent K(+) efflux; higher ones also produced a small influx of protons, probably through a K(+)/H(+) exchanger, resulting in a decrease of the internal pH of the cells and the efflux of material absorbing at 260nm and phosphate. The cell membrane was not permeabilized. The K(+) efflux with miconazole was dependent directly on the medium pH. This efflux results in an increased membrane potential, responsible for an increased Ca(2+) uptake and other effects. These effects were not observed with two triazolic antifungals. A decrease of the Zeta (ζ) potential was observed at low concentrations of miconazole. Although the main effect of these antifungals is the inhibition of ergosterol synthesis, K(+) efflux is an important additional effect to be considered in their therapeutic use. Under certain conditions, the use of single mutants of several transporters involved in the movements of K(+) allowed to identify the participation of several antiporters in the efflux of the cation.     

Involvement of glutathione peroxidase 1 in growth and peroxisome formation in Saccharomyces cerevisiae in oleic acid medium

Saccharomyces cerevisiae is able to use some fatty acids, such as oleic acid, as a sole source of carbon. β-oxidation, which occurs in a single membrane-enveloped organelle or peroxisome, is responsible for the assimilation of fatty acids. In S. cerevisiae, β-oxidation occurs only in peroxisomes, and H(2)O(2) is generated during this fatty acid-metabolizing pathway. S. cerevisiae has three GPX genes (GPX1, GPX2, and GPX3) encoding atypical 2-Cys peroxiredoxins. Here we show that expression of GPX1 was induced in medium containing oleic acid as a carbon source in an Msn2/Msn4-dependent manner. We found that Gpx1 was located in the peroxisomal matrix. The peroxisomal Gpx1 showed peroxidase activity using thioredoxin or glutathione as a reducing power. Peroxisome biogenesis was induced when cells were cultured with oleic acid. Peroxisome biogenesis was impaired in gpx1? cells, and subsequently, the growth of gpx1? cells was lowered in oleic acid-containing medium. Gpx1 contains six cysteine residues. Of the cysteine-substituted mutants of Gpx1, Gpx1(C36S) was not able to restore growth and peroxisome formation in oleic acid-containing medium, therefore, redox regulation of Gpx1 seems to be involved in the mechanism of peroxisome formation.     

Involvement of the Saccharomyces cerevisiae hydrolase Ldh1p in lipid homeostasis

Here, we report the functional characterization of the newly identified lipid droplet hydrolase Ldh1p. Recombinant Ldh1p exhibits esterase and triacylglycerol lipase activities. Mutation of the serine in the hydrolase/lipase motif GXSXG completely abolished esterase activity. Ldh1p is required for the maintenance of a steady-state level of the nonpolar and polar lipids of lipid droplets. A characteristic feature of the Saccharomyces cerevisiae Δldh1 strain is the appearance of giant lipid droplets and an excessive accumulation of nonpolar lipids and phospholipids upon growth on medium containing oleic acid as a sole carbon source. Ldh1p is thought to play a role in maintaining the lipid homeostasis in yeast by regulating both phospholipid and nonpolar lipid levels.     

Accumulation of magnesium ions during fermentative metabolism in Saccharomyces cerevisiae

When cells of Saccharomyces cerevisiae were grown aerobically under glucose-repressed conditions, ethanol production displayed a hyperbolic relationship over a limited range of magnesium concentrations up to around 0.5 mM. A similar relationship existed between available Mg²⁺ and ethanol yield, but over a narrower range of Mg²⁺ concentrations. Cellular demand for Mg²⁺ during fermentation was reflected in the accumulation patterns of Mg²⁺ by yeast cells from the growth medium. Entry of cells into the stationary growth phase and the time of maximum ethanol and minimum sugar concentration correlated with a period of maximum Mg²⁺ transport by yeast cells. The timing of Mg²⁺ transport fluxes by S. cerevisiae is potentially useful when conditioning yeast seed inocula prior to alcohol fermentations. Received 04 March 1996/ Accepted in revised form 21 August 1996     

Accumulation and intracellular compartmentation of lithium ions in Saccharomyces cerevisiae

Abstract Accumulation of Li⁺ in Saccharomyces cerevisiae X2180-1B occured via an apparent stoichiometric relationship of 1: 1 (K⁺/Li⁺) when S. cerevisiae was incubated in the presence of 5 and 10 mM LiCl for 3 h. Other cellular cations (Mg²⁺, Ca²⁺ and Na⁺) did not vary on Li⁺ accumulation, although lithium chemistry dictates a degree of similarity to Group I and II metal cations. Compartmentation of Li⁺ was mainly in the vacuole which accounted for 85% of the Li⁺ accumulated after a 6-h incubation period. The remainder was located in the cytosol with negligible amounts being bound to cell fragments including the cell wall. Transmission electron microscopy of Li⁺-loaded cells revealed enlarged vacuoles compared with control cells. This asymmetric cellular distribution may therefore enhance tolerance of S. cerevisiae to Li⁺ and ensure that essential metabolic processes in the cytosol are not disrupted.     

酵母交配过程的极性生长及其调控机制

酿酒酵母单倍体细胞能够与相反交配型的单倍体细胞发生交配。交配时酿酒酵母放弃原有出芽位点,根据信息素的浓度梯度,重新选择生长位点,向相反交配型细胞伸出突起进行极性生长。交配因子受体指导选择交配突起的位点,通过G蛋白激活Ste20p,将信号经由Ste11p、Ste7p和Fus3p组成的MAPK模块传递到Far1p和Ste12p等因子,调控相关基因的转录,抑制原有的出芽位点,选择新的生长位点,并使细胞周期停止在G1期,G蛋白与Cdc24p、Cdc42p和Bem1p等蛋白作用,聚集在细胞,使得肌协蛋白细胞骨架在交配突起处聚集,呈极性化分布,使细胞发生极性生长。